Activation of Autophagy Restores Nitric Oxide as the Primary Mechanism of Flow‐Mediated Dilation in the Absence of Telomerase Reverse Transcriptase Activity

Abstract

Arteriolar vasodilation to shear stress, termed flow‐mediated dilation, is endothelium‐dependent, primarily mediated by nitric oxide (NO) in health, and hydrogen peroxide (H2O2) in subjects with coronary artery disease (CAD). Previous evidence has demonstrated that autophagy and the catalytic subunit of telomerase, telomerase reverse transcriptase (TERT), govern the mediator of arteriolar FMD such that inhibition of telomerase activity or autophagy initiate a switch in mechanism of dilation from NO to H2O2 in non‐CAD, while activation of either telomerase or autophagy switch the mechanism of dilation from H2O2 to NO in CAD arterioles. We examined whether autophagy is necessary for the vascular effects of TERT utilizing a rat loss of function model that phenocopies the vascular changes observed in subjects with CAD. We hypothesized that activation of autophagy will switch the mechanism of dilation from H2O2 to NO, despite absence of TERT. Trehalose, an autophagy activator was administered in drinking water (2%) for 4 weeks. Mesenteric arterioles (3rd or 4th order) were isolated from TERT KO and wild‐type (WT) littermates and prepared for video and fluorescent microscopy. Control KO and WT animals received normal drinking water. Internal diameters were measured following increases in intraluminal flow (pressure gradient) in the presence/absence of a NOS inhibitor (L‐NAME; 100μM) or H2O2 scavenger (Peg‐Catalase, Peg‐Cat; 500 units/mL). Shear‐induced autophagy was examined using Lysotracker Red DND‐99 in response to sustained shear (100 cmH2O) for 1 hour. FMD is expressed as % maximal dilation, while Lysotracker data is expressed as relative fold change in fluorescent signal above static. In arterioles from untreated KO rats H2O2 is the primary mechanism of FMD (Vehicle: 64±9% vs. L‐NAME: 70±8% vs. Peg‐Cat: 16±4%; n=9; P<0.05), while NO predominates in WT arterioles (P<0.05). 4 weeks of 2% trehalose supplementation initiated a switch to NO as the primary mechanism of dilation in KO arterioles (Vehicle: 55±9% vs. L‐NAME: 19±6% vs. Peg‐Cat: 58±13%; n= 6–8; P<0.05). Similarly, in WT arterioles, NO is maintained as the primary mechanism of dilation (P<0.05). In KO rats shearinduced autophagy was reduced, while trehalose augmented the shear‐induced activation of autophagy in both KO and WT relative to untreated KO and WT animals (P<0.05). Our data demonstrate that trehalose initiates a switch from H2O2 to NO in KO rats, and augments shearinduced autophagy in both KO and WT. Taken together, autophagy is necessary for maintenance of NO as the primary mechanism of FMD in the absence of TERT activity.Support or Funding InformationFunded by T32GM089586‐09 (WEH), R01‐HL135901‐02 (DDG), and R01‐HL133029‐02 (AMB)